Stress affects virtually everyone in society at some point in life and it has major deleterious effects on health. It contributes to numerous pathologies, including disorders in the cardiovascular, immune and reproductive systems, as well as sleep disorders, depression and substance abuse. In spite of this, the neuroendocrine mechanisms regulating the physiological responses to stress remain poorly understood, particularly the regulation and role of prolactin secretion. The overall goal of these studies is to understand the regulation of prolactin secretion during stress and its impact on the response of hypothalamic-pituitary adrenal (HPA) axis to stress. The major hypothesis is that the endogenous opiate, Orphanin FQ/Nociceptin (OFQ/N), is an important neuropeptide regulating prolactin secretion during stress. The OFQ/N knock out model will be used because it is ideally suited to these studies. First, knock out mice do not respond to stress in the same way as wild type mice; this is true for both males and females. Acute stress increased plasma prolactin levels in wild type, but not knock out males. Also, OFQ/N knock out, but not wild type females continued to respond to repeated stress, suggesting PRL feedback is not active when OFQ/N is absent. The role and mechanisms of action of OFQ/N in mediating these gender differences will be examined in Specific Aims 1 and 2. Second, knock out post-partum females are not hyperprolactinemic and do not seem to have increased PRL-R mRNA in the choroid plexus of the brain, making them much different from normal post-partum females. In Specific Aim 3, we will take advantage of this difference to examine the role and mechanisms involved in OFQ/N regulation of hyperprolactinemia and prolactin's role and mechanisms of action in mediating attenuation of the HPA axis response to stress. Plasma corticosterone levels, a reliable indicator of stress activation, and plasma PRL levels will be measured. To determine the mechanisms regulating the stress response, tuberoinfundibular dopaminergic neuronal activity, anterior pituitary prolactin content, prolactin receptor expression in the choroid plexus and paraventricular nucleus (PVN) of the hypothalamus and CRH mRNA levels in the PVN will be determined. Specificity of action will be demonstrated by administering prolactin antiserum to block effects mediated by increased prolactin levels. Although little is known about the neuroendocrine effects of OFQ/N, our studies indicate that OFQ/N plays an important role in prolactin regulation. These studies will determine the mechanisms and physiological significance of OFQ/N in mediating the prolactin response to stress and the effects of prolactin on HPA axis activation. Understanding the neuroendocrine mechanisms regulating the stress response will lead to improved treatments for alleviating stress and anxiety, and hence, have a positive impact on health. Stress affects virtually everyone in society at some point in life, and has major deleterious effects. It contributes to numerous pathologies, including, but not limited to, disorders in the cardiovascular, immune and reproductive systems, as well as sleep disorders, depression and substance abuse. The cost to society in terms of job productivity and health care is staggering. The focus of these studies is on the neuroendocrine response to stress, particularly on the role of the endogenous opiate, Orphanin FQ/Nociceptin, a purported anxiolytic, in prolactin regulation. Prolactin is critically important in the stress response, but its mechanisms of action are poorly understood. Understanding the neuroendocrine regulation of stress will lead to improved treatments to alleviate stress and anxiety, and hence, have a positive impact on health. [unreadable] [unreadable] [unreadable]